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Complete Genome Sequence of Fish Pathogen Aeromonas hydrophila AL06-06 Hasan C. Tekedar,a Attila Karsi,a Ali Akgul,a Safak Kalindamar,a Geoffrey C. Waldbieser,b Tad Sonstegard,c Steven G. Schroeder,c Mark L. Lawrencea College of Veterinary Medicine, Mississippi State University, Mississippi State, Mississippi, USAa; Warmwater Aquaculture Research Unit, Agriculture Research Service, United States Department of Agriculture, Stoneville, Mississippi, USAb; Bovine Functional Genomics Laboratory, Agricultural Research Service, United States Department of Agriculture, Beltsville, Maryland, USAc
Received 12 March 2015 Accepted 17 March 2015 Published 23 April 2015 Citation Tekedar HC, Karsi A, Akgul A, Kalindamar S, Waldbieser GC, Sonstegard T, Schroeder SG, Lawrence ML. 2015. Complete genome sequence of fish pathogen Aeromonas hydrophila AL06-06. Genome Announc 3(2):e00368-15. doi:10.1128/genomeA.00368-15. Copyright © 2015 Tekedar et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 3.0 Unported license. Address correspondence to Mark L. Lawrence, [email protected].
A
eromonas species are Gram-negative facultative anaerobes that have worldwide distribution in aquatic environments (1, 2), and they can be isolated from domesticated pets, invertebrate species, birds, ticks, insects, and natural soils (3). Also, many aeromonads cause disease in fish. Aeromonas hydrophila is a motile species that has primarily been considered an opportunistic pathogen in fish and humans (3). We previously reported the complete genome sequence of A. hydrophila ML09-119 (1), which represents a clonal group of A. hydrophila strains causing outbreaks of bacterial septicemia in commercial channel catfish aquaculture in the southeastern United States. We now report the complete genome sequence of A. hydrophila AL06-06, which was isolated from a diseased goldfish in 2006 from the Auburn University Southeastern Cooperative Fish Disease Laboratory in Greensboro, Alabama. The AL06-06 genome sequence will be used for comparative genomic purposes with other sequenced Aeromonas strains, particularly those causing disease in catfish. The genome of A. hydrohila AL06-06 was fully sequenced using an Illumina Genome Analyzer IIx (a total of 6,629,874 reads, with 150⫻ coverage). Trimming, error correction, contig creation, and quality control of sequence reads were conducted using CLC Workbench 6.5.1 (CLC Bio) and Sequencher 5.2.3 (Gene Codes Corporation). De novo assembly was performed by CLC Workbench 6.5.1. Scaffolded gaps were closed by Sanger sequencing of PCR amplicons. For the unscaffolded gaps, single-primer PCR was used for amplification of sequence templates (4). Ribosomal operons and other repetitive regions were amplified and completely resequenced to create a reliable assembly. NCBI’s Prokaryotic Genomes Automatic Annotation Pipeline (PGAAP) (5) was used for AL06-06 genome annotation, followed by submission to GenBank. The complete genome of A. hydrophila genome comprises 4,884,823 bp with 61.3% G⫹C content. It contains 4,453 predicted genes, of which 4,235 are protein coding. A total of 112 tRNAs and 10 rRNA operons were predicted by PGAAP.
March/April 2015 Volume 3 Issue 2 e00368-15
The A. hydrophila AL06-06 genome was also annotated by RAST (6) to facilitate comparison with the A. hydrophila ML09119 genome. Based on functional comparative results, A. hydrophila AL06-06 has 81 unique elements including arsenic resistance genes, heme and hemin uptake-utilization systems, some membrane transport genes for type I and type V secretion systems, transposable elements, and nitrogen metabolism genes. In particular, strain AL06-06 has a specific arsenic resistance mechanism that is missing in the A. hydrophila ML09-119 genome. Due to its ubiquitous distribution in the environment, A. hydrophila is prone to arsenic exposure (7). The A. hydrophila AL06-06 genome also has three plasmids compared to strain ML09-119, which does not carry any plasmids. In summary, the complete genome of A. hydrophila AL06-06 contributes to our knowledge of A. hydrophila virulence and environmental adaptations, and it is especially useful for comparison with other fish- and human-pathogenic A. hydrophila strains. Nucleotide sequence accession numbers. The completed genome sequence of A. hydrophila AL06-06 was deposited in GenBank under the accession no. CP010947 (the version described in this paper is CP010947.1, GI:764079125). The accession numbers for the plasmids are CP010948 (for pAH06-06-1; the version described in this paper is version CP010948.1, GI: 764083361), CP010949 (for pAH06-06-2; the version described in this paper is version CP010949.1, GI:764083365), and CP010950 (for pAH06-06-3; the version described in this paper is version CP010950.1, GI:764083373). ACKNOWLEDGMENTS This work was supported by USDA NIFA grant 2013-67015-21313, Mississippi State University College of Veterinary Medicine, and the USDA Agricultural Research Service CRIS project 6402-31000-009-00D. We thank Michelle Banes and Stephen Reichley for their technical assistance.
Genome Announcements
genomea.asm.org 1
Downloaded from http://genomea.asm.org/ on May 25, 2015 by NDSU
Aeromonas hydrophila occurs in freshwater environments and infects fish and mammals. Here, we report the complete genome sequence of Aeromonas hydrophila AL06-06, which was isolated from diseased goldfish and is being used for comparative genomic studies with A. hydrophila strains that cause bacterial septicemia in channel catfish aquaculture.
Tekedar et al.
REFERENCES 1. Tekedar HC, Waldbieser GC, Karsi A, Liles MR, Griffin MJ, Vamenta S, Sonstegard T, Hossain M, Schroeder SG, Khoo L, Lawrence ML. 2013. Complete genome sequence of a channel catfish epidemic isolate, Aeromonas hydrophila strain ML09-119. Genome Announc 1(5):e00755-13. http:// dx.doi.org/10.1128/genomeA.00755-13. 2. Woo PTK, Bruno DW (ed). 2011. Fish diseases and disorders: 3. Viral, bacterial and fungal infections. CAB International, Oxfordshire, United Kingdom. 3. Janda JM, Abbott SL. 2010. The genus Aeromonas: taxonomy, pathogenicity, and infection. Clin Microbiol Rev 23:35–73. http://dx.doi.org/10.1128/ CMR.00039-09. 4. Karlyshev AV, Pallen MJ, Wren BW. 2000. Single-primer PCR procedure for rapid identification of transposon insertion sites. Biotechniques 28: 1078, 1080, 1082.
5. Angiuoli SV, Gussman A, Klimke W, Cochrane G, Field D, Garrity G, Kodira CD, Kyrpides N, Madupu R, Markowitz V, Tatusova T, Thomson N, White O. 2008. Toward an online repository of standard operating procedures (SOPs) for (meta)genomic annotation. Omics J Integr Biol 12: 137–141. http://dx.doi.org/10.1089/omi.2008.0017. 6. Overbeek R, Olson R, Pusch GD, Olsen GJ, Davis JJ, Disz T, Edwards RA, Gerdes S, Parrello B, Shukla M, Vonstein V, Wattam AR, Xia F, Stevens R. 2014. The SEED and the rapid annotation of microbial genomes using subsystems technology (RAST). Nucleic Acids Res 42:D206 –D214. http://dx.doi.org/10.1093/nar/gkt1226. 7. Goswami R, Ghosh D, Saha DR, Padhy PK, Mazumder S. 2011. Effect of acute and chronic arsenic exposure on growth, structure and virulence of Aeromonas hydrophila isolated from fish. Microb Pathog 50:63– 69. http:// dx.doi.org/10.1016/j.micpath.2010.10.005.
Downloaded from http://genomea.asm.org/ on May 25, 2015 by NDSU
2 genomea.asm.org
Genome Announcements
March/April 2015 Volume 3 Issue 2 e00368-15
Complete Genome Sequence of Fish Pathogen Aeromonas hydrophila AL06-06 Hasan C. Tekedar,a Attila Karsi,a Ali Akgul,a Safak Kalindamar,a Geoffrey C. Waldbieser,b Tad Sonstegard,c Steven G. Schroeder,c Mark L. Lawrencea College of Veterinary Medicine, Mississippi State University, Mississippi State, Mississippi, USAa; Warmwater Aquaculture Research Unit, Agriculture Research Service, United States Department of Agriculture, Stoneville, Mississippi, USAb; Bovine Functional Genomics Laboratory, Agricultural Research Service, United States Department of Agriculture, Beltsville, Maryland, USAc
Received 12 March 2015 Accepted 17 March 2015 Published 23 April 2015 Citation Tekedar HC, Karsi A, Akgul A, Kalindamar S, Waldbieser GC, Sonstegard T, Schroeder SG, Lawrence ML. 2015. Complete genome sequence of fish pathogen Aeromonas hydrophila AL06-06. Genome Announc 3(2):e00368-15. doi:10.1128/genomeA.00368-15. Copyright © 2015 Tekedar et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 3.0 Unported license. Address correspondence to Mark L. Lawrence, [email protected].
A
eromonas species are Gram-negative facultative anaerobes that have worldwide distribution in aquatic environments (1, 2), and they can be isolated from domesticated pets, invertebrate species, birds, ticks, insects, and natural soils (3). Also, many aeromonads cause disease in fish. Aeromonas hydrophila is a motile species that has primarily been considered an opportunistic pathogen in fish and humans (3). We previously reported the complete genome sequence of A. hydrophila ML09-119 (1), which represents a clonal group of A. hydrophila strains causing outbreaks of bacterial septicemia in commercial channel catfish aquaculture in the southeastern United States. We now report the complete genome sequence of A. hydrophila AL06-06, which was isolated from a diseased goldfish in 2006 from the Auburn University Southeastern Cooperative Fish Disease Laboratory in Greensboro, Alabama. The AL06-06 genome sequence will be used for comparative genomic purposes with other sequenced Aeromonas strains, particularly those causing disease in catfish. The genome of A. hydrohila AL06-06 was fully sequenced using an Illumina Genome Analyzer IIx (a total of 6,629,874 reads, with 150⫻ coverage). Trimming, error correction, contig creation, and quality control of sequence reads were conducted using CLC Workbench 6.5.1 (CLC Bio) and Sequencher 5.2.3 (Gene Codes Corporation). De novo assembly was performed by CLC Workbench 6.5.1. Scaffolded gaps were closed by Sanger sequencing of PCR amplicons. For the unscaffolded gaps, single-primer PCR was used for amplification of sequence templates (4). Ribosomal operons and other repetitive regions were amplified and completely resequenced to create a reliable assembly. NCBI’s Prokaryotic Genomes Automatic Annotation Pipeline (PGAAP) (5) was used for AL06-06 genome annotation, followed by submission to GenBank. The complete genome of A. hydrophila genome comprises 4,884,823 bp with 61.3% G⫹C content. It contains 4,453 predicted genes, of which 4,235 are protein coding. A total of 112 tRNAs and 10 rRNA operons were predicted by PGAAP.
March/April 2015 Volume 3 Issue 2 e00368-15
The A. hydrophila AL06-06 genome was also annotated by RAST (6) to facilitate comparison with the A. hydrophila ML09119 genome. Based on functional comparative results, A. hydrophila AL06-06 has 81 unique elements including arsenic resistance genes, heme and hemin uptake-utilization systems, some membrane transport genes for type I and type V secretion systems, transposable elements, and nitrogen metabolism genes. In particular, strain AL06-06 has a specific arsenic resistance mechanism that is missing in the A. hydrophila ML09-119 genome. Due to its ubiquitous distribution in the environment, A. hydrophila is prone to arsenic exposure (7). The A. hydrophila AL06-06 genome also has three plasmids compared to strain ML09-119, which does not carry any plasmids. In summary, the complete genome of A. hydrophila AL06-06 contributes to our knowledge of A. hydrophila virulence and environmental adaptations, and it is especially useful for comparison with other fish- and human-pathogenic A. hydrophila strains. Nucleotide sequence accession numbers. The completed genome sequence of A. hydrophila AL06-06 was deposited in GenBank under the accession no. CP010947 (the version described in this paper is CP010947.1, GI:764079125). The accession numbers for the plasmids are CP010948 (for pAH06-06-1; the version described in this paper is version CP010948.1, GI: 764083361), CP010949 (for pAH06-06-2; the version described in this paper is version CP010949.1, GI:764083365), and CP010950 (for pAH06-06-3; the version described in this paper is version CP010950.1, GI:764083373). ACKNOWLEDGMENTS This work was supported by USDA NIFA grant 2013-67015-21313, Mississippi State University College of Veterinary Medicine, and the USDA Agricultural Research Service CRIS project 6402-31000-009-00D. We thank Michelle Banes and Stephen Reichley for their technical assistance.
Genome Announcements
genomea.asm.org 1
Downloaded from http://genomea.asm.org/ on May 25, 2015 by NDSU
Aeromonas hydrophila occurs in freshwater environments and infects fish and mammals. Here, we report the complete genome sequence of Aeromonas hydrophila AL06-06, which was isolated from diseased goldfish and is being used for comparative genomic studies with A. hydrophila strains that cause bacterial septicemia in channel catfish aquaculture.
Tekedar et al.
REFERENCES 1. Tekedar HC, Waldbieser GC, Karsi A, Liles MR, Griffin MJ, Vamenta S, Sonstegard T, Hossain M, Schroeder SG, Khoo L, Lawrence ML. 2013. Complete genome sequence of a channel catfish epidemic isolate, Aeromonas hydrophila strain ML09-119. Genome Announc 1(5):e00755-13. http:// dx.doi.org/10.1128/genomeA.00755-13. 2. Woo PTK, Bruno DW (ed). 2011. Fish diseases and disorders: 3. Viral, bacterial and fungal infections. CAB International, Oxfordshire, United Kingdom. 3. Janda JM, Abbott SL. 2010. The genus Aeromonas: taxonomy, pathogenicity, and infection. Clin Microbiol Rev 23:35–73. http://dx.doi.org/10.1128/ CMR.00039-09. 4. Karlyshev AV, Pallen MJ, Wren BW. 2000. Single-primer PCR procedure for rapid identification of transposon insertion sites. Biotechniques 28: 1078, 1080, 1082.
5. Angiuoli SV, Gussman A, Klimke W, Cochrane G, Field D, Garrity G, Kodira CD, Kyrpides N, Madupu R, Markowitz V, Tatusova T, Thomson N, White O. 2008. Toward an online repository of standard operating procedures (SOPs) for (meta)genomic annotation. Omics J Integr Biol 12: 137–141. http://dx.doi.org/10.1089/omi.2008.0017. 6. Overbeek R, Olson R, Pusch GD, Olsen GJ, Davis JJ, Disz T, Edwards RA, Gerdes S, Parrello B, Shukla M, Vonstein V, Wattam AR, Xia F, Stevens R. 2014. The SEED and the rapid annotation of microbial genomes using subsystems technology (RAST). Nucleic Acids Res 42:D206 –D214. http://dx.doi.org/10.1093/nar/gkt1226. 7. Goswami R, Ghosh D, Saha DR, Padhy PK, Mazumder S. 2011. Effect of acute and chronic arsenic exposure on growth, structure and virulence of Aeromonas hydrophila isolated from fish. Microb Pathog 50:63– 69. http:// dx.doi.org/10.1016/j.micpath.2010.10.005.
Downloaded from http://genomea.asm.org/ on May 25, 2015 by NDSU
2 genomea.asm.org
Genome Announcements
March/April 2015 Volume 3 Issue 2 e00368-15
